Hepatitis B virus infections are one of the leading causes of chronic hepatitis. Infection results in a broad range of clinical symptoms from mild, nonapparent disease to fulminant hepatitis to hepatocellular carcinoma. Due to lack of in vitro infection system for HBV, the events of infectious processes are poorly understood. HBV encodes a regulatory protein termed HBx. While many functions have been attributed to HBx, a clear picture of how this protein participates in establishing infectious process has not emerged. In this competing renewal grant application, the focus of our study will be on the detailed characterization of HBx s association with mitochondria and exploring the functional consequences of that association. First, we propose to identify the mitochondrial targeting domain within HBx protein. Using those mutants, which fail to associate with mitochondria, further characterization of various possible functions of HBx within mitochondria will be investigated. These include, the ability of HBx via its interaction with the outer membrane channel VDAC to alter mitochondrial membrane potential (delta-psim), generation of reactive oxygen species (ROS). Ca+2 homeostasis. and permeability transition among others. Alteration of these functions will be correlated with induction of gene expression via NF-kB, AP-1, NF-AT and STAT-3 transcription factors. The results of these studies will delineate the molecular mechanisms in the induction of HBV-induced liver disease pathogenesis including hepatocellular carcinoma.